Studies in this group have focused on what controls the organization and distribution of organelles and transport intermediates within the endomembrane system of higher eucaryotes. This has led to an investigation of the characteristics of the membrane pathways between specific organelles and the role of microtubules and microtubule- associated proteins in facilitating membrane transport. Focusing primarily in the ER and Golgi apparatus, which together function in the generation, maintenance and sorting of membrane components within the endomembrane system, we have made three major findings: (1) Delivery of membrane from the ER into the Golgi apparatus in fibroblast cell lines occurs through distinct transport intermediates which arise from multiple peripheral exit sites within the ER and then move toward the centrally located Golgi apparatus in a microtubule-facilitated process; (2) Er to Golgi transport intermediates exist as tubular-vesicular structures which are largely devoid of resident ER and golgi components but are enriched in the integral membrane protein, p53, the peripheral "coat" protein, bCOP, and the microtubule plus-end directed motor protein, kinesin; and (3) The predominant localization of membrane-bound kinesin on pre-Golgi structures appears to represent a steady-state distribution, with kinesin constitively cycling within the ER/Golgi system (as has been previously shown for p53). Based on these observations, we have proposed that kinesin acts as a pus-end directed motor protein in the recycling of membrane from the Golgi to the ER ad in intra-ER membrane movements (both of which involve microtubule plus end directed movements), and that kinesin associated with membrane structures moving from the ER toward the Golgi complex (i.e. toward the minus ends of microtubules) has either inhibited or altered motor activity. An in vitro approach is currently being used to investigate what factors might regulate kinesin motor activity in the ER/Golgi membrane system.